There is no reproducible technique for establishing donor specific tolerance in clinical transplantation. Dr. Madsen and his colleagues in the MGH Transplantation Biology Research Center (TBRC) have established a large animal model of cardiac allograft tolerance by co transplanting a kidney and heart allograft under a short course of cyclosporine A (CyA) in class IMHC disparate inbred miniature swine. Heart/kidney co transplantation provides the only consistent and reproducible model of nonablative cardiac allograft tolerance in large animals. However, the cell populations involved in the induction and maintenance of tolerance in this model have not been elucidated. Recent studies at the TBRC have indicated that the donor kidney and host thymus play a critical role in tolerant heart/kidney recipients. Based on this work, we hypothesize that host thymus-derived regulatory T cells, generated by cells or antigens derived from the donor kidney, are responsible for the induction of tolerance to cardiac allografts in heart/kidney recipients. We will test this hypothesis by performing a longitudinal in vitro characterization of regulatory T cells from different lymphoid compartments in tolerant heart/kidney recipients.